Manganese proteins and enzymes are important in a diverse range of biological functions. The relative amounts and distribution of Mn in humans is known, however, due to the relatively weak binding of Mn, the mechanisms by which Mn can affect human health are only now beginning to be unraveled. Manganese plays an essential role in many cellular processes of both eukaryotes and prokaryotes. Two mononuclear Mn enzymes (Dioxygenase and Acetone Carboxylase) will be investigated in this proposal due to their unique mechanisms of substrate activation. In addition, metal transport and regulation systems are a growing area of study. Recently, a Mn transport regulatory protein (MntR) has been structurally characterized to show a dinuclear Mn site. A spectroscopic characterization of the Mn centers in MntR and how it recognizes DNA in response to Mn will be investigated. A fundamental advance in the interpretation of EPR spectra for both mono- and dinuclear Mn centers will allow an unprecedented ability to quantitatively characterize the active Mn sites of proteins and enzymes. This will be applied to the above proteins, and also for a characterization of the dinuclear Mn enzyme Bacteriophage lambda-Phosphatase. Insight into the mechanism of oxygen and nitrogen activation by Mn and Fe enzymes will be derived from studies of biomimetic complexes. The detection and characterization of intermediates in such mechanisms is critical for an understanding of the mechanism. Two different families of biomimetic complexes and their oxygen or nitrogen derived intermediates will be characterized.